Affiliation: School of Physiology and Pharmacology, University of Bristol Bristol, UK.

ABSTRACTOur goal was to develop and refine a decerebrate arterially perfused rat (DAPR) preparation that allows the complete bladder filling and voiding cycle to be investigated without some of the restrictions inherent with in vivo experimentation [e.g., ease and speed of set up (30 min), control over the extracellular milieu and free of anesthetic agents]. Both spontaneous (naturalistic bladder filling from ureters) and evoked (in response to intravesical infusion) voids were routinely and reproducibly observed which had similar pressure characteristics. The DAPR allows the simultaneous measurement of bladder intra-luminal pressure, external urinary sphincter-electromyogram (EUS-EMG), pelvic afferent nerve activity, pudendal motor activity, and permits excellent visualization of the entire lower urinary tract, during typical rat filling and voiding responses. The voiding responses were modulated or eliminated by interventions at a number of levels including at the afferent terminal fields (intravesical capsaicin sensitization-desensitization), autonomic (ganglion blockade with hexamethonium), and somatic motor (vecuronium block of the EUS) outflow and required intact brainstem/hindbrain-spinal coordination (as demonstrated by sequential hindbrain transections). Both innocuous (e.g., perineal stimulation) and nociceptive (tail/paw pinch) somatic stimuli elicited an increase in EUS-EMG indicating intact sensory feedback loops. Spontaneous non-micturition contractions were observed between fluid infusions at a frequency and amplitude of 1.4 ± 0.9 per minute and 1.4 ± 0.3 mmHg, respectively and their amplitude increased when autonomic control was compromised. In conclusion, the DAPR is a tractable and useful model for the study of neural bladder control showing intact afferent signaling, spinal and hindbrain co-ordination and efferent control over the lower urinary tract end organs and can be extended to study bladder pathologies and trial novel treatments.

Figure 5: Non-micturition contractions (NMCs). (A) Low amplitude NMCs occurred under basal conditions, when bladder volume was low. Each NMC was accompanied by tonic firing of the EUS. (B) During fluid infusion in the same preparation, NMCs became larger in amplitude with bladder distension until voiding was triggered. (C) When brainstem control had deteriorated (as indicated by a loss of phrenic activity and voiding), the NMCs became biphasic and their amplitude significantly increased. In the first contractile phase of each NMC, tonic EUS firing was observed. The subsequent single burst of the EUS was followed by a second pressure oscillation (dotted line) and they could now be associated with leakage of fluid.

Mentions:
Spontaneous NMCs were seen as asymmetrical bladder wall movements in most preparations, even when the bladder volume was low (e.g., immediately post-void or when infusion was stopped). The NMCs had a frequency of 1.4 ± 0.9 per minute and intra-luminal pressure increase of 1.4 ± 0.3 mmHg (n = 12; Figure 5A). The NMCs were associated with phasic increases in EUS–EMG activity that presumably maintained continence. During active filling, as bladder volume increased, the amplitude of NMCs also increased (although the basal bladder pressure remained low), until void (Figure 5B). When brainstem function had deteriorated, marked by weakening phrenic nerve activity and loss of voiding, NMC amplitude increased significantly [5.8 ± 0.4 mmHg; (n = 12) paired t-test P < 0.0001; Figure 5C] but not frequency. In addition, marked changes were seen in the pressure waveform of NMCs after loss of brainstem control where biphasic waves were commonly seen (Figure 5C). Associated EUS activity was still present, but leakage of small volumes of urine was frequently seen but this was not a coordinated void.

Figure 5: Non-micturition contractions (NMCs). (A) Low amplitude NMCs occurred under basal conditions, when bladder volume was low. Each NMC was accompanied by tonic firing of the EUS. (B) During fluid infusion in the same preparation, NMCs became larger in amplitude with bladder distension until voiding was triggered. (C) When brainstem control had deteriorated (as indicated by a loss of phrenic activity and voiding), the NMCs became biphasic and their amplitude significantly increased. In the first contractile phase of each NMC, tonic EUS firing was observed. The subsequent single burst of the EUS was followed by a second pressure oscillation (dotted line) and they could now be associated with leakage of fluid.

Mentions:
Spontaneous NMCs were seen as asymmetrical bladder wall movements in most preparations, even when the bladder volume was low (e.g., immediately post-void or when infusion was stopped). The NMCs had a frequency of 1.4 ± 0.9 per minute and intra-luminal pressure increase of 1.4 ± 0.3 mmHg (n = 12; Figure 5A). The NMCs were associated with phasic increases in EUS–EMG activity that presumably maintained continence. During active filling, as bladder volume increased, the amplitude of NMCs also increased (although the basal bladder pressure remained low), until void (Figure 5B). When brainstem function had deteriorated, marked by weakening phrenic nerve activity and loss of voiding, NMC amplitude increased significantly [5.8 ± 0.4 mmHg; (n = 12) paired t-test P < 0.0001; Figure 5C] but not frequency. In addition, marked changes were seen in the pressure waveform of NMCs after loss of brainstem control where biphasic waves were commonly seen (Figure 5C). Associated EUS activity was still present, but leakage of small volumes of urine was frequently seen but this was not a coordinated void.

Affiliation:
School of Physiology and Pharmacology, University of Bristol Bristol, UK.

ABSTRACTOur goal was to develop and refine a decerebrate arterially perfused rat (DAPR) preparation that allows the complete bladder filling and voiding cycle to be investigated without some of the restrictions inherent with in vivo experimentation [e.g., ease and speed of set up (30 min), control over the extracellular milieu and free of anesthetic agents]. Both spontaneous (naturalistic bladder filling from ureters) and evoked (in response to intravesical infusion) voids were routinely and reproducibly observed which had similar pressure characteristics. The DAPR allows the simultaneous measurement of bladder intra-luminal pressure, external urinary sphincter-electromyogram (EUS-EMG), pelvic afferent nerve activity, pudendal motor activity, and permits excellent visualization of the entire lower urinary tract, during typical rat filling and voiding responses. The voiding responses were modulated or eliminated by interventions at a number of levels including at the afferent terminal fields (intravesical capsaicin sensitization-desensitization), autonomic (ganglion blockade with hexamethonium), and somatic motor (vecuronium block of the EUS) outflow and required intact brainstem/hindbrain-spinal coordination (as demonstrated by sequential hindbrain transections). Both innocuous (e.g., perineal stimulation) and nociceptive (tail/paw pinch) somatic stimuli elicited an increase in EUS-EMG indicating intact sensory feedback loops. Spontaneous non-micturition contractions were observed between fluid infusions at a frequency and amplitude of 1.4 ± 0.9 per minute and 1.4 ± 0.3 mmHg, respectively and their amplitude increased when autonomic control was compromised. In conclusion, the DAPR is a tractable and useful model for the study of neural bladder control showing intact afferent signaling, spinal and hindbrain co-ordination and efferent control over the lower urinary tract end organs and can be extended to study bladder pathologies and trial novel treatments.